Coryneform bacteria are microorganisms used to produce various chemical materials used in many applications, such as animal feed, medicines, and food including L-lysine, L-threonine, and various nucleic acids. A strain of coryneform bacteria showing high productivity can be developed through genetic engineering and metabolic engineering. To obtain such a strain of coryneform bacteria showing high productivity, a gene relating to various metabolic pathways needs to be expressed in the coryneform bacteria. To this end, a suitable promoter must be developed.
Generally, in coryneform bacteria, a gene is expressed under a promoter inherently included therein. (see, for example, Journal of Bacteriology, 181(19), 6188-6191, 1999). Meanwhile, the structure of a promoter sequence for expressing a gene in coryneform bacteria is not known, whereas the structures of other industrial microorganisms, such as E. coli and Bacillus subtilis, are known. Therefore, the following method has been suggested to produce promoters enabling the expression of a gene in coryneform bacteria. First, a promoter region of a gene that is resistant to an antibiotic, such as chloramphenicol, is removed. Separately, a chromosomal DNA separated from coryneform bacteria is cleaved using a suitable restriction enzyme, and the resulting fragment is introduced to the gene from which the promoter region is removed. Then, the obtained gene is used to transform coryneform bacteria to produce a transformed strain and the antibiotic resistance of the transformed strain is measured: (see Gene, 102, 93-98, 1991; Microbiology, 142, 1297-1309, 1996.) In particular, a very small number of promoters used in Corynebacterium ammoniagenesis, a known nucleic acid producing microorganism, has been developed. For example, a promoter having about 10% higher activity than a tac promoter is used in E. coli (see Biotechnol. Lett. 25, 1311-1316, 2003.) However, when it is used in a mass expression of genes, such a promoter exhibits low efficiency. U.S. Pat. No. 5,593,781 discloses a promoter DNA which is separated from a Brevibacterium flavum strain MJ-233 (FERM BP-1497) and has higher activity than a tac promoter. However, such a promoter DNA that is separated from a Brevibacterium genus may not be operable in other bacteria. Therefore, there is a need to develop a promoter sequence that is derived from commercially available Corynebacterium ammoniagenes, and has high activity in other bacteria.
Accordingly, the inventors of the present invention searched for a strong promoter sequence in Corynebacterium ammoniagenes and found that a promoter according to the present invention can express genes with high activity in Corynebacterium ammoniagenes. 